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Biochemical and physiological changes associated with estrogenic activity in Xenopus laevis : a model for the detection of endocrine disruption

Hurter, Etienne (2003-03)

Dissertation (PhD)--University of Stellenbosch, 2003.

Thesis

ENGLISH ABSTRACT: Concern has increased that widespread adverse effects are occurring in humans and wildlife
populations as a result of exposure to environmental chemicals (mostly man-made) that
disrupt the normal functioning of the endocrine system. Many pharmaceutical, agricultural
and industrial chemicals, acting as endocrine modulating compounds (EDCs), have been
detected in an accumulated form in food, in drinking water and in the environment.
Although the levels of these chemicals can be measured analytically, it is important to
evaluate biological activity. For this, animal models are used and relevant bioassays
developed. These assays are based on biological markers, which are specific
xenobiotically-induced physiological responses and are usually deviations in cellular or
biochemical components, processes or structures. Vitellogenin is a large protein complex,
produced in the liver under estrogen control and circulates in the plasma, destined for
incorporation into the developing oocyte in female oviparous vertebrates. Since
vitellogenin production is low or nonexistent in males, its presence may therefore be
interpreted as evidence of exposure to estrogenic environmental contaminants. In this study
the African Clawed Frog, Xenopus laevis was used as model to study the effects of
estrogens on biochemical and physiological parameters associated with vitellogenesis. As a
starting point the seasonal female reproductive cycle in a natural Xenopus laevis population
in terms of ovarian state, plasma vitellogenin and plasma estrogen levels was studied. It
was shown that plasma vitellogenin and estrogen levels were seasonal and correlated to a
seasonal ovarian cycle, which peaked during spring. However, although seasonality
existed, there were reproductively active individuals at any time during the year. Increases
in plasma estrogen levels predated increases in plasma vitellogenin levels and ovarian maturation. Lipoprotein profiles, as well as plasma cholesterol, triglyceride and
phospholipid concentrations were determined and it was shown that estrogen affected these
in such a way that these parameters could be used as additional markers in estrogen
contamination studies. In order to develop an in vitro bioassay to screen for estrogenic
activity, the use of hepatic tissue cultures was investigated. Optimal culture conditions were
established and increased sensitivity in the estrogenic response was obtained by using liver
slices from male frogs that were pre-treated with estrogen. Validation studies proved that
this bioassay could be employed for routine screening of water and chemical samples. In
order to refine the Xenopus laevis vitellogenin ELISA and liver slice bioassay, existing
polyclonal anti-vitellogenin antibodies were replaced by culturing monoclonal antibodies.
Selected antibodies were characterised and ELISAs developed and validated. This study
showed that the newly developed Xenopus laevis vitellogenin ELISA and liver slice
bioassay have the potential to be employed in environmental monitoring programmes.

ENGLISH ABSTRACT: Many pesticides have shown health impacts in wildlife and humans, including endocrine system modulation. The potential health effects of the pesticides elicit concerns considering high volume being used ...

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African clawed frogs (Xenopus laevis) are often referred to as ‘purely aquatic’ but there are many publications which suggest extensive overland movements. Previous reviews which considered the topic have not answered the ...

Due to both deliberate and accidental introductions, invasive African Clawed Frog (Xenopus laevis) populations have become established worldwide. In this study, we investigate the geographic origins of invasive X. laevis ...

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A sensitive ex vivo bioassay for detecting environmental estrogens and estrogen mimics was developed using Xenopus liver slice culture. Vitellogenin synthesis by these cultures was used as biomarker for estrogenic activity ...

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Terrestrial isopods have been used in a number of studies as test organisms concerning the effects of chemicals in a (eco-)toxicological context. They are saprophages, which are in direct contact with polluted plant material. ...